Cell culture isolation of Hepatitis E Virus Genotype 3 Strain obtained from human feces

• Published in: Acta biológica colombiana Vol. 24; no. 3; pp. 503 - 508
• Main Authors: Quintero-Gil, Carolina, Mirazo, Santiago, Parra-Suescún, Jaime, López-Herrera, Albeiro, Mainardi, Victoria, Arbiza, Juan, Orduz, Sergio
• Format: Journal Article
• Language: English
• Published: Universidad Nacional de Colombia, Facultad de Ciencias, Departamento de Biología 01.09.2019; Universidad Nacional de Colombia
• Subjects: A549 cells; BIOLOGY; hepevirus; isolation; RNA virus; Aislamiento; virus ARN; A549 cells; RNA virus; isolation; células A549; hepevirus
• ISSN: 0120-548X; 1900-1649
• DOI: 10.15446/abc.v24n3.79351

Hepatitis E virus (HEV) is considered one of the leading causes of acute viral hepatitis worldwide, and about 20 million infections and approximately 57 000 deaths occurred every year. However, little is known about the replicative virus cycle due to the absence of a consensus cell culture model. A549 cell line is considered susceptible to HEV genotype 3, however, both viral strain and cell culture conditions could affect the viral isolation in vitro. The objective of this work was to isolate in vitro an HEV-3 strain obtained from human feces. To this, a genotype 3 HEV strain previously identified by genetic characterization was inoculated in A549 monolayers, and incubated for two hours at 37 °C. Five days post-infection, cells were passaged (subcultured) for the first time, and serial passages were done on average every four days during 41 days. HEV replication was evaluated through RT-qPCR in each passage, and reinfection of the cell line with the viral progeny derived from A549 infected monolayers was assessed through immunofluorescence and RT-qPCR. Viral RNA was detected in each passage from infected monolayers, and the highest amount was found after 26 days (2 x 106 copies/μL). In reinfection assay, capsid antigen was detected perinuclearly and forming foci, and 1x104 copies/μL of viral RNA was detected after 96 hours post infection. This shows that HEV recovered from the cell lysate monolayers was infectious. This viral isolate offers a critical tool to study the unknown aspect of HEV infection.